Clostridium perfringens enterotoxin (CPE) is emerging as one of the most important bacterial virulence factors for gastrointestinal (G.I.) disease. There is compelling epidemiologic evidence for the involvement of CPE in one of the most common human food poisonings; recent studies also link CPE-positive C. perfringens to other serious human gastrointestinal illnesses and to Sudden Infant Death Syndrome. CPE has a unique action on mammalian membranes which is distinguishable from all other known toxins. Every step of CPE action involves close co-participation between CPE and mammalian membrane proteins. The long-term objective of this project is to understand the role of CPE (and CPE-positive isolates) in human disease; these studies may ultimately identify strategies to prevent or treat CPE-associated illnesses. To accomplish this objective, the following specific aims are proposed: 1) to test a recent hypothesis that a 50kDa membrane protein serves as a functional CPE receptor (the binding of CPE to the CPE receptor is essential for cytotoxicity) using "anti-receptor antibody" CPE binding inhibition studies and cloning of the CPE receptor, 2) to further clarify CPE action by a) determining the topologic orientation of CPE in membranes and b) dissecting individual events in CPE action using both biochemical and ultrastructural techniques, 3) to high-resolution map CPE functional regions (and eventually combine this information with CPE structural data) to identify how the CPE molecule participates in cytotoxicity and 4) to dissect the molecular pathogenesis of CPE-positive isolates by studying the importance of CPE (and possibly other toxins) in the virulence of these isolates. These important studies are expected to i) increase the knowledge of how CPE participates in disease, ii) broaden perspectives of pathophysiologic mechanisms of intestinal diseases, iii) allow use of CPE as a defined probe of plasma membrane structure/function relationships and iv) provide a model for understanding sporulation-associated virulence factor regulation in the biomedically important genus Clostridia. These findings should have significance for numerous biomedical disciplines seeking to understand intestinal physiology under normal and disease conditions and how enteric pathogens cause G.I. disease.